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Conceptually simple yet efficient cell labeling techniques offering broader applicability are warranted despite many methods requiring intricate construction procedures. One such technique is ADAR (adenosine deaminase acting on RNA)-mediated RNA sensing, a live cell labeling technique that is based on the expression and abundance of cell-type-specific RNAs. Here, we utilized the optimized version, CellREADR (Cell access through RNA sensing by Endogenous ADAR), to establish a feasible tracing system for mouse spermatogonial stem cells (mSSCs) which are refractory to CRISPR-based reporter gene knock-in. We identified several previously unreported features of CellREADR, including RNA interference induced by double-strand RNA formation that is obligatorily generated during the normal operation of the CellREADR system. More importantly, we established its application for long-term labeling of mSSCs and monitoring mSSC differentiation induced by retinoic acid (RA) treatment in vitro. This work offers a practical solution for dynamic monitoring of mSSC self-renewal and differentiation and supports that CellREADR can be developed into more versatile and efficient tools in stem cell research.<\/jats:p>","DOI":"10.3390\/biotech15040077","type":"journal-article","created":{"date-parts":[[2026,9,8]],"date-time":"2026-09-08T11:23:51Z","timestamp":1788866631000},"page":"77","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Long-Term Labeling and Differentiation Monitoring of Mouse Spermatogonial Stem Cells Using CellREADR"],"prefix":"10.3390","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0009-0005-2869-5814","authenticated-orcid":false,"given":"Shuaitao","family":"Hu","sequence":"first","affiliation":[{"name":"State Key Laboratory of Organ Regeneration and Reconstruction, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China"},{"name":"Institute for Stem Cell and Regeneration, Chinese Academy of Sciences, Beijing 100101, China"},{"name":"Beijing Institute for Stem Cell and Regenerative Medicine, Beijing 100101, China"},{"name":"University of Chinese Academy of Sciences, Beijing 100049, China"},{"name":"Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650204, China"}],"role":[{"vocabulary":"crossref","role":"author"},{"role":"conceptualization","vocabulary":"credit"},{"role":"investigation","vocabulary":"credit"},{"role":"methodology","vocabulary":"credit"},{"role":"formal-analysis","vocabulary":"credit"},{"role":"writing-original-draft","vocabulary":"credit"},{"role":"visualization","vocabulary":"credit"}]},{"given":"Lin","family":"Yan","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Organ Regeneration and Reconstruction, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China"},{"name":"Institute for Stem Cell and Regeneration, Chinese Academy of Sciences, Beijing 100101, 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